HPLC测定帕金森模型小鼠脑中D-丝氨酸的含量

Determination of D-serine in Brain of Parkinson's Disease Mouse by High Performance Liquid Chromatography

以邻苯二甲醛（OPA）-N-异丁酰基-D-半胱氨酸（IBDC）为手性衍生试剂，醋酸钠-甲醇-乙腈混合液为流动相，采用梯度洗脱,荧光检测，建立了测定生物样品中微量D-丝氨酸（D—Ser）的高效液相色谱法。Set对映体在20min内完全分离。D—Ser在30～1500nmol／L时，其浓度与峰面积呈良好线性关系（R=0．9995）。当信噪比为3（S／N=3）时，D—Ser的检出限为10nmol／L。将方法运用于帕金森病（Parkinson’s disease，PD）模型小鼠中脑及大脑皮质中D—Ser含量的测定，结果表明，模型组脑组织中D—Set的含量与对照组相比，整体呈显著下降趋势，推测D—Ser可能参与了PD的发病机制。

A reversed high performance liquid chromatography （HPLC） method was established for the determination of the trace amounts of D-serine （D-Ser） in biological samples. Ser enantiomers were precolumn-derivatized with o-phthalaldehyde （OPA） and N-isobutyryl-D-cysteine （IBDC）. The Separation of the amino acid enantiomers was achieved on a Damonsil^TM C18 column using a gradient solvent program and the column eluent was monitored using fluorescence detection. The limit of detection （LOD） of 10 nmol/L was reached when signal of noise ratio was 3. A linear range was found between peak area and concentration in the range of 30 to 1 500 nmol/L for D-Ser. The levels of D-Ser in midbrains and brain cortex of PD mouse were detected by the present method. Compared with the controls,the level of D-Ser was obviously decreased in brains of PD mouse. The result suggested that D-Ser might participate in PD pathogenesis.